1. Field of the Invention
The present invention relates generally to the fields of molecular toxicology, biochemistry and molecular biology. More specifically, the present invention relates to the isolation and cloning of the human ARSA-I gene.
2. Description of the Related Art
Little is known about mammalian detoxification systems involved with environmental heavy metal salt toxins. Some chemotherapeutic drugs are heavy metal-based and the development of resistance to such drugs occurs frequently during treatment and constitutes a major obstacle to the cure of even sensitive tumors. Resistance is thought to be due to the selection for and overgrowth of drug-resistant cells that arise through spontaneous somatic mutation. Biochemical studies have not succeeded in conclusively identifying the basis of resistance, but they have defined several mechanisms which can contribute to resistance. Platinum drugs do not participate in the multidrug resistance phenotype conferred by either the mdrl (P-glycoprotein) or M R P genes and only the ATP-dependent glutathione GS-X pump is known to modulate resistance to cisplatin.
Several transport-protein complexes that mediate the detoxification of heavy metal salts have been identified in bacteria and yeast, and appear to be structurally and functionally conserved throughout evolution. Resistance to arsenite, antimonite, tellurite, and arsenate in E. coli. and S. aureus is mediated by the plasmid-borne ars operon whose gene products together form an ATP-dependent pump that extrudes oxyanions and results in decreased uptake of these metalloid compounds.
Heavy metal salts are toxic to many types of organisms, and are important industrial toxins for man. Resistance to heavy metal salts in bacteria is mediated by specific plasmid-borne multicomponent ATP-dependent efflux systems (1,2). In E. coli, resistance to arsenite, arsenate and antimonite is mediated by the well-characterized ars operon (3) that contains two regulatory (arsR and arsD) and three structural genes (arsA, B and C) (2,4).
The gene arsA codes for an oxyanion-dependent ATPase that associates with the product of the arsB gene which is a putative channel-forming transmembrane protein. The ATP-binding cassette (ABC) of the arsA protein belongs to a superfamily of genes with a modified NTP-binding motif that is distinct from that present in other ATPases including the cation-translocating transporters (5,6). The arsA gene codes for a 583 amino acid (63 kDa) catalytic subunit with two ATP-binding cassette domains, whereas the arsB gene codes for a 429 amino acid (45.5 kDa) inner membrane protein with 12 transmembrane spanning domains that is postulated to serve both as the anion channel and an anchor for the ArsA protein (7). Together these two proteins transport arsenite and antimonite out of the cell. The arsC gene codes for a 141 amino acid (16 kDa) reductase capable of utilizing GSH as a cofactor to reduce arsenate (As.sup.+5) to arsenite (As.sup.+3), thus making it a substrate for ars transport system (8).
The prior art is deficient in the lack of the isolation and cloning of the human ARSA-I gene. The present invention fulfills this longstanding need and desire in the art.